Cell shape change and motility are critical components in a wide range of biological processes in mammals, including embryonic development, tissue repair, angiogenesis, and immune system function. Cell shape change and motility also are involved in pathological events, such as cancer metastasis. While progress has been made in identifying components of signal transduction pathways leading to cell motility, a complete model of the mechanism is still lacking. The precise roles of many proteins implicated in these pathways are not yet elucidated, and a number of mechanisms that cells use for movement may exist.
Cell motility is dependent on regulated actin filament assembly, rearrangement, and disassembly. A large and growing number of proteins are known to regulate and modulate the state of the actin cytoskeleton, and some appear to have partly overlapping functions. For example, actin polymerization and filament assembly can be accomplished through de novo nucleation of new filaments by the Arp2/3 complex or through elongation of existing filaments at free barbed or fast-growing ends, generated by filament severing and/or regulated dissociation of bound barbed-end capping proteins (uncapping). Similarly, multiple routes exist for actin filament bundling, crosslinking, and disassembly (depolymerization).
In addition to these end-point mechanisms of actin dynamics, a number of different upstream signaling pathways leading to changes in the actin cytoskeleton and cell morphology and behavior have become apparent. The small Ras-related GTPases, e.g., Rac, Rho, and Cdc42, in particular, have been implicated in the regulation of the actin cytoskeleton and cell shape, and each plays a distinct and specialized role. Rho proteins are associated generally with formation of contractile actin/myosin bundles, stress fibers, and focal adhesions. Rac proteins particularly are associated with formation of lamellipodia (broad, sheet-like membrane protrusions at the leading edge in the direction of movement). Lamellipodial cell crawling resulting from activation of Rac proteins is considered to be the most prevalent form of animal cell motility. Cdc42 particularly is most associated with formation of filpodia (finger-like membrane protrusions) and the control of cell polarity. The Rho family small GTPases also have roles in other cellular processes, such as control of cell growth and cell-cell adhesion. In addition to these small GTPases, phosphoinositides and calcium are known to regulate actin dynamics and cell migration. However, a comprehensive understanding of the signaling cascades leading to cell motility and the relationship between these regulators remains elusive.
Progress in the art would be facilitated by the availability of effective inhibitors of cell motility. A number of compounds that target actin directly exist. The best known compounds are the cytochalasins, which are cell-permeable destabilizers of actin filaments, and phalloidin, which is a cell-impermeable stabilizer of actin filaments (J. A. Cooper, J. Cell Biol, 105 (1987)). In addition, latrunculins are cell-permeable disrupters of actin filaments (I. Spector, Science, 219, 493 (1983)). Jasplakinolide is a cell-permeable stabilizer of actin filaments (M. R. Bubb et al., Chem., 269, 14869 (1994)). A few compounds that target proteins upstream of the actin cytoskeleton are known, such as the Rho-kinase inhibitor Y-27632 (M. Uehata et al., Nature, 389, 990 (1997), and myosin light chain kinase inhibitors, such as ML-g (M. Saitoh et al., Biochem. Biophys, Res. Commun., 140, 280 (1986)). Recently, a cyclic peptide dimer was discovered that inhibits the activity of N-WASP, a protein involved in Cdc42-mediated actin nucleation by the Arp2/3 complex (J. R. Peterson et al., Proc. Natl. Acad. Sci. USA, 98, 10624 (2001)). Nevertheless, there is a dearth of available cell-permeable compounds that affect actin dynamics and cell motility by inhibiting specific components of signaling pathways to the actin cytoskeleton.
Presently, very few specific inhibitors of cell motility are available, even though a great potential exists for such drugs as a complement to existing anticancer therapies. Cell shape change and motility are involved at two rate-limiting steps in cancer progression: angiogenesis (i.e., blood vessel recruitment) and metastasis (i.e., spreading of a tumor from one location in the body to other locations). In combination with cell growth inhibitors, treatment with specific cell motility inhibitors has the potential to provide a more efficacious treatment of a cancer, analogous to the multiple drug approach for treatment of HIV infection and AIDS.
In particular, cell motility inhibitors have potential uses such as, but not limited to, (a) an anticancer drug targeting angiogenesis, (b) an anticancer drug targeting metastasis, and (c) an anticancer drug targeting cell growth.